Many Gram-negative pathogens use the type III pathway to transport protein toxins across the bacterial cell envelope. Pathogenic Yersinia spp., Y. pestis, Y. pseudotuberculosis and Y. enterocolitica, secrete 14 proteins (Yops, Yersinia outer proteins) via the type III pathway. During infection, substrate recognition and transport are regulated in a manner that allows the Yersinia type III pathway to direct Yops to the extracellular medium or into the cytosol of host cells. Together the type III mechanisms are essential to provide for bacterial escape from the host's innate immune response and to allow for microbial multiplication in human lymphoid tissues. Although many tissue culture cells serve as targets for type III injection, only some, but certainly not all cells of an infected host are injected by Yersinia. Yersinia lacking the surface adhesins YadA and invasin (InvA) fail to bind and inject tissue culture cells, suggesting that binding of bacterial ligands to surface receptors of host cells is a prerequisite for the type III injection of Yop proteins. During infection, Yersiniae perceive host-specific environmental cues that are transduced to relieve repression of the type III pathway. Although several regulatory mechanisms of gene expression and type Ill transport have emerged, the genes and signaling pathways that activate type III secretion are not yet established. Preliminary work suggests that the type III machinery may recognize its substrates via signals encoded in yop mRNA. These secretion signals, suggested to reside in the first seven to fifteen codons of yop mRNA, presumably act by promoting transcript specific codon recognition and/or binding of factors that couple ribosomal protein synthesis to type III secretion. Work proposed herein focuses on characterizing the elements, factors and mechanisms of substrate recognition and regulation of type III transport. Other work is aimed at characterizing the mechanism of Yersinia target cell recognition. Together these studies will enhance our understanding of the molecular mechanisms that control the establishment of Yersinia infection and may provide insight into future strategies for prevention or therapy of these bacterial diseases. [unreadable] [unreadable]